Organ procurement currently poses one of the major problems in organ transplantation, as the number of patients requiring transplants far exceeds the number of organs available. A path for eliminating the shortage of donor organs is to develop the technologies required to transplant nonhuman organs into humans, i.e., xenotransplantation.
Potential sources of xenogeneic organs include nonhuman primates and pigs. There are serious problems with considering nonhuman primates as donors. Chimpanzees, the closest nonhuman primates phylogenetically, are far too rare to be considered. Baboons are too small to be an appropriate donor for most organ transplants. Even the largest baboons weigh less than 40 kg. In addition, the gestation times and productivity of primates would not allow a commercially significant generation of source animals.
The physiology of many organ systems of pigs has been shown to be highly similar to the human counterparts (Sachs, D. H. 1994. Veterinary Immunology & Immunopathology, 43:185-191). Pigs come in many different sizes including the breed described as miniature porcine. The miniature swine have a variety of advantages as potential xenograft donors. They achieve adult weights of approximately 100-150 kg, a size which is more compatible to human weights than the domestic pig which reaches weights of over 500 kg. Through a selective breeding program over the past 20 years partially inbred, miniature swine have been produced (Sachs et al. 1976. Transplantation 22: 559-567; Sachs, 1992. In Swine as models in biomedical research. M. Swindle, D. Moody, and L. Phillips, eds. (Ames Iowa State Univ. Press) pp 3-15 ; Sachs, 1994. veterinary Immunology & Immunopathology, 43: 185-191).
These breeding program has resulted in herds of animals that are genetically well characterized. These animals have been used in large animal model studies for many years and have, like their domestic counterparts, very favorable breeding characteristics for the production of donor animals.
A central concern regarding xenotransplantation is the risk of xenosis, infection by organisms transferred with the xenograft into both the transplant recipient and the general population. The risk of viral infection is increased in transplant recipients by the presence of factors commonly associated with viral activation, e.g., immune suppression, graft-versus-host disease, graft rejection, viral co-infection, and cytotoxic therapies. Type C retroviruses from cells of swine origin have been characterized (Arida, E. and Hultin, T. 1977. Am. J. Public Health 67: 380; Armstrong et al. 1971. J. Gen. Virol. 10: 195-198; Benveniste, R. E. and Todaro, G. J. 1973. Proc. Natl. Acad. Sci. USA 70:3316-3320; Bouillant et al. 1975. J. Gen. Virol. 27: 173-180; Frazier, M. E. 1985. Arch. Virol. 83: 83-97; Lieber et al. 1975. Virology 66:616-619; Suzuka et al. 1985. FEBS Lett. 183: 124-128; Suzuka et al. 1986. FEBS Lett. 198: 339-343; Todaro et al. 1974. Virology 58: 65-74; Woods et al. 1973. J. Virol. 12: 1184-1186; Akiyoshi et al. 1988 J. Virol. 72: 4503-4507); as yet, no disease following infection by these viruses has been identified. A recent report demonstrated that a virus from the PK15 (porcine kidney-derived) cells can infect human cells in vitro (Patience et al., 1997. Nature Medicine 3:276-282).
Characterization of swine cells and cell lines has resulted in the identification of at least three pig endogenous retrovirus sequences (PERV-A, -B, -C), (WO 97/40167; WO 97/21836; Le Tissier, et al. 1997. Nature 389: 681-682; Czauderna, F. et al. 1998: Genbank Accession Number Y17013). These sequences have distinct envelope (env) genes but share highly conserved sequences in the rest of the genome. Southern blot analysis of pig tissues and cell lines (Patience et al., 1997. Nature Medicine 3:276-282) showed the presence of numerous loci in DNA extracted from normal pig hearts and from pig cell lines. The banding profile for normal pig hearts is similar to that of the pig cell lines and is typical of an endogenous inherited retrovirus suggesting heterogeneity with approximately 50 integration sites. These results were confirmed and extended to analysis of MHC-inbred miniature swine where the numbers of potentially full-length provirus copies are approximately 8 to 15 per genome for inbred and outbred swine and 10 to 20 in PK15 cells (Akiyoshi et al. 1998. J. Virology, 72:4503-4507).
The env gene product determines host range and cell tropism. Host range analyses using retrovirus vectors bearing corresponding env proteins showed that PERV-A and PERV-B envs have wider host range including several human cell lines compared to PERV-C env which infected only 2 pig cell lines and 1 human cell line. All three classes of PERVs have been shown to infect pig cells. Receptors for PERV-A and PERV-B have been shown to be present on cells of some other species, including mink, rat, mouse and dog. Interference studies showed that the three PERV strains use distinct receptors to each other and to a number of other type C mammalian retroviruses.